In the biotechnical field, fluorescent dyes are routinely used as sensitive, non-isotopic labels. These labels are used to identify and locate a variety of cell structures, ranging from malignant tumors to specific chromosomes in a DNA sequence. A variety of devices have been designed to read fluorescent-labeled samples.
Gel electrophoresis is one technique commonly used in conjunction with fluorescent dyes and other markers to identify specific molecules as well as other tagged units. In this technique an electric field is used to cause the migration of the tagged units through a gel or other solution.
In U.S. Pat. No. 4,874,492 a gel electrophoresis system is disclosed in which samples are treated with fluorescent markers prior to applying them to an electrophoretic gel. The gel is illuminated with a UV source and the fluorescence pattern is detected with a cooled charge-coupled-device (CCD) two-dimensional detector array. The CCD array is cooled to at least -25 degrees C. in order to improve light sensitivity and increase the dynamic range.
In U.S. Pat. No. 5,162,654 a system is disclosed to optically determine which of four fluorophores is fluorescing in an electrophoresis gel. Fluorescence emitted by the gel passes first through four separate band pass filters and then through four wedge prisms. As a result of this optical configuration, the emitted fluorescence is imaged on four discrete areas on the detector array. The specific fluorophore exited by the irradiation source is determined by comparing the relative intensities of the fluorescence detected in the four detection areas.
In U.S. Pat. No. 5,294,323 the disclosed gel electrophoresis system utilizes a vertical electrophoresis plate. A laser beam passes horizontally through the gel in a direction perpendicular to the longitudinal axis of the electrophoresis plate. The emitted fluorescence is reflected to a solid state imaging sensor such that the reflected pattern is parallel to the direction of the laser beam.
In U.S. Pat. No. 5,324,401 a fluorescence detection system for capillary electrophoresis is disclosed which provides for the simultaneous excitation and detection of fluorescent probes within a plurality of capillaries. The excitation source is a laser which is coupled to the capillaries through an optical fiber bundle. The fluorescence from the capillary array is focussed through a lens and imaged onto a CCD camera for analysis.
In a paper by Sutherland et al. entitled "Electronic Imaging System for Direct and Rapid Quantitation of Fluorescence from Electrophoretic Gels: Application to Ethidium Bromide-Stained DNA" published in Analytical Biochemistry 163, 446-457 (1987), the authors describe an imaging system which uses a CCD camera. The CCD camera quantifies the fluorescence received from electrophoretic gels, chromatograms, and other sources. The paper describes several sources of non-uniformities which impact the ability of the system to obtain accurate results.
From the foregoing, it is apparent that an improved electrophoresis apparatus is desired which enables accurate quantitative fluorescence measurements.